We have recently documented that hypoxia, one of the major etiologic factors of vaso-occlusive crisis (VOC) and acute chest syndrome (ACS), markedly increase red blood cell (RBC)-endothelial cell (EC) adhesion and that the predominant receptor combination involved in this effect is the VCAM-1-VLA-4 pairing (Blood 88:2311-2320, 1996). In preliminary studies, we now show that exposure of human umbilical vein ECs (HUVECs) to oleic acid (OA), a fatty acid (FA) reported to be elevated in ACS, not only stimulated basal sickle RBC adherence, but also enhanced the hypoxia- induced response. On the other hand, docosahexaenoic acid (DHA) (a FA present in fish oil) inhibited the adhesion process. In aim I, we propose to strengthen these preliminary data by examining the effects of OA and DHA on adherence of sickle RBCs to HUVECs. We will also study human lung microvascular ECs (HLMECs). We will initially delineate the receptors and the mechanism involved. Moreover, the protective effects of DHA, if confirmed, could potentially lead to therapeutic interventional clinical strategies. Aim II seeks to exploit further our previous findings that hypoxia mediates its effect on RBC-Ec adherence via up-regulation of VCAM- 1. Our premise is that in ACS massive pulmonary adhesion occurs as a result of hypoxia-induced VCAM-1 up-regulation which is not counteracted by the normal cyto-protective mediators including NO and/or prostacyclin (PGI/2). We hypothesize that during ACS the levels of cyto-protective agents that potentially down-regulate the expression of VCAM-1 are decreased, whereas those inflammatory mediators that up-regulate VCAM-1 expression are increased. The cyto-protective agents produced by endothelium and cells in the local microenvironment that we have selected to investigate in patients during episodes of ACS include NO, prostacyclin (PGI/2) and 13-HODE. NO studies will be done in expired air, white cells, and urine. The inflammatory mediators involved in induction of VCAM-1 expression that we will also study in ACS include the cytokineses (IL-1 and TNF-alpha) hydroxyeicosanoids (15-HETE/HPETE, 12-HETE and LTB/4), PAF, TxA/2 and endothelin-1. The cyto-protective effects of NO and PGI/2 will also be evaluated in vitro in HUVECs and subsequently in HLVMECs where VCAM-1 up-regulation will be initiated by hypoxia. Since inflammatory mediators released by the circulating cellular elements of blood have particular relevance to ACS, we will also explore the cyto-protective effects of NO and PGI/2 in vitro against the activation induced by these pathogenic factors. Our preliminary studies on the cyto-protector and cytotoxic effectors demonstrate that the latter up regulated VCAM-1 and that the former do protect against VCAM-1 up-regulation. Thus, our in vitro and in vivo studies related to ACS should provide us with the information necessary to assess whether the rationale for potential interventional strategies (i.e. NO and/or PGI/2) is sound such that they can subsequently be translated into therapeutic reality.